User manual

Basic requirements

Metabiome was built under different bioinformatics considerations to simplify its use. Prior to installation you require:

  1. Linux terminal. If you are a windows user, you can use a virtual machine. For more information, please check VirtualBox.

  2. CONDA as a package manager. We recommend installing MINICONDA as it contains all necessary packages, and the latest version can be downloaded from this page. To install it, run the following commands:

bash Miniconda3-latest-Linux-x86_64.sh

  1. Access to a computer server, given the large size of metagenomic data sets and the complexity of subsequent analyzes.

Installation

In order to install, clone Metabiome into your machine and execute install.sh by running the following commands:

git clone -b master https://github.com/Nesper94/metabiome.git
cd Metabiome/
bash install.sh

You can also download Metabiome directly from the Github page.

The installation script will do the following:

  • Create a link to the Metabiome’s main script in ~/.local/bin/ and if necessary add this folder to PATH so that you can use Metabiome from whatever location in your terminal.

  • Create Conda environments with all the software you can use with Metabiome.

  • Install the _metabiome bash completion script to make Metabiome’s commands easier to use.

Uninstalling

If for some reason you need to uninstall Metabiome, all you have to do is go to the Metabiome’s folder and execute the uninstall.sh script:

cd scripts/
bash uninstall.sh

Additional command line options

Some Metabiome commands accept the flag -opts, if this flag is used then it must be the last flag used in the command. After this flag you can write options from the native program executed by Metabiome. For example, if you want to assemble a large and complex metagenome like soil and need to use the flag --presets from MEGAHIT, then you would do the following:

metabiome megahit -i in_dir -o out_dir -opts --presets meta-large

Naming convention

Metabiome’s working needs that the input files agree with one of the two most widespread naming conventions for paired end sequences: Illumina _R1_/_R2_ naming convention and _1/_2 naming convention.

File extensions accepted are:

  • .fq

  • .fastq

  • .fq.gz

  • .fastq.gz

Metabiome modules

Metabiome contains 9 modules that comprise the necessary tools for the analysis of the main points within metagenomics. They are separated by conda environments, created from a .yaml file, which describes the software that each one implements and the required version. These files are stored in the conda_envs/ directory. A module can have one or more than one tool or software, and each one has a separate script to execute it. They are stored in the scripts/ directory.

The following is the list of software contained in each one of the Metabiome modules:

metabiome-preprocessing

  • Bowtie 2 (v2.3)

  • FastQC (v0.11)

  • MultiQC (v1.6)

  • Trimmomatic (v0.39)

metabiome-taxonomic-profiling

  • MetaPhlAn3 (v3.0)

  • HUMAnN3 (v3.0)

metabiome-taxonomic-binning

  • Kaiju (v1.7)

  • Kraken2 (v2.1)

  • Krona (v2.7)

metabiome-extract16S

  • BBMap (v38.87)

metabiome-genome-assembly

  • MEGAHIT (v1.2)

  • QUAST (v5.0)

  • SPAdes (v3.12)

metabiome-concoct

  • CONCOCT (v1.1)

  • CoverM (v0.6)

metabiome-maxbin2

  • MaxBin2 (v2.2)

metabiome-metabat2

  • MetaBAT2 (v2.15)

metabiome-das_tool

  • DAS Tool (v1.1.2)

Debug mode

Sometimes errors can appear when you try to run a command. If you are a developer and/or have knowledge in bash, you can use the Metabiome’s debug mode to obtain more information about the error. The debug mode is turned on by setting the DEBUG_METABIOME environment variable:

export DEBUG_METABIOME="yes"

Then you can run the command again and see a detailed output that can help in the debugging process.

Software native help message

Metabiome commmand line option includes an additional flag named -hh that allows the users to see the programs native help message. For example, if you want more information about the assembler metaSPAdes the -hh would show the help message from this software:

metabiome metaspades -hh

Note

It seems that Krona doesn’t have a help command, and FastQC and MultiQC usage in our pipeline is very simple, so they don’t have this option.